The human glucocorticoid receptor (GR) is expressed in almost all cells of the body. Upon binding of glucocorticoid hormones such as cortisol the receptor is translocated to the cell nucleus and activates a tissue-specific set of target genes. The fact that GR target genes vary from one tissue to another results in a pleiotropic pattern of GR effects in different tissues.
Many of the physiological actions of glucocorticoid hormones are of medical interest and present potential areas for clinical intervention. For example, in Cushing's syndrome, excess GR activity leads to high blood pressure. In the brain, abnormalities in the GR pathway have been linked to depression and mood disorders; and, in the lung, such abnormalities have been associated with asthma and chronic airway diseases.
One of the best-characterized clinical activities of glucocorticoid hormones is their anti-inflammatory action, which is due to their immuno-suppressive effects. Exposure of T-cells to glucocorticoid hormones leads to T-cell anergy and interferes with T-cell activation. For a recent review, see Rhen, T. et al. (2005). N. Engl. J. Med. 353(16):1711-23. Long-term treatment with glucocorticoids leads to serious side effects like diabetes and osteoporosis. See discussion in Rosen, J. et al. (2005) Endocr. Rev. 26(3):452-64. Moreover, suppression of the entire immune system can lead to the reactivation of latent viruses (see Reinke, P. et al. (1999) Transpl Infect Dis 1(3): 157-64) and interferes with immunotherapy approaches; e.g., the delivery of a beneficial subset of immune cells to patients.
Many of the problems associated with the GR overactivation that accompanies glucocorticoid treatment could be solved if a method was available which allows selective disruption of GR function in a subset of cells; e.g., a characterized population of T-cells. One such method would be to alter the sequence of the gene encoding the GR. Indeed, the ability to manipulate (i.e. edit) the DNA sequence at specific locations in the genome has been a major goal of human genome biology for some time. A variety of techniques have previously been tested for this purpose, but the frequencies of genome modification achieved with these methods have generally been too low for therapeutic applications. See, e.g., Yanez, R. J. et al. (1998) Gene Ther. 5(2): 149-159.
Another important application of genome editing is the insertion of clinically useful transgenes into the genome. However, a crucial requirement for any genome editing method is that it allow for targeted insertion into a defined location. The importance of the requirement for precisely targeted integration of a therapeutic transgene was underscored by the recent observation, in a clinical trial for treatment of X-linked SCID that the random integration of transgenes used for human gene therapy resulted, in certain cases, in insertional mutagenesis which led to oncogenic transformation of target cells. Hacein-Bey-Abina, S. et al. (2003). Science 302(5644):415-9.
Various methods and compositions for targeted cleavage of genomic DNA have been described. Such targeted cleavage events can be used, for example, to induce targeted mutagenesis, induce targeted deletions of cellular DNA sequences, and facilitate targeted recombination at a predetermined chromosomal locus. See, for example, United States patent application publications US 2003/0232410 (Dec. 18, 2003), US 2005/0026157 (Feb. 3, 2005), US 2005/0064474 (Mar. 24, 2005), US 2005/0208489 (Sep. 22, 2005) and US 2006/0188987 (Aug. 24, 2006); the disclosures of which are incorporated by reference in their entireties for all purposes. Targeted integration of exogenous sequences can also be accomplished. See U.S. patent application Ser. No. 11/493,423 (filed Jul. 26, 2006) the disclosure of which is incorporated by reference in its entirety for all purposes. See also PCT WO 2005/084190 (Sep. 15, 2005), the disclosure of which is incorporated by reference in its entirety for all purposes.
However, methods and compositions for specific cleavage of the human glucocorticoid receptor gene, and for modulation of immune function by modification of the GR gene, have not heretofore been described.